Staining for viability testing,germination and maturation of Sambucus nigra L. pollen in vitro

Freshly released pollen of black elderberry (Sambucus nigra L.) was incubated under various culture conditions until germination was achieved. Optimal conditions for germination were determined and used for maturation of unicellular microspores in vitro. Staining with 5-diphenyltetrazolium bromide, propidium iodide and iodine potassium iodide was used to assess pollen viability, nuclear phase and maturation, respectively. The germination rate was highest when fresh pollen was agitated at 40 rpm in Petri dishes containing a liquid medium consisting of Brewbaker and Kwack salts, 15% (w/v) sucrose, 500 mg/l MES sodium salt, at pH 5.0; germination reached nearly 70% after only 1 h in culture. Under these conditions, and with addition of 200 mg/l glutamine, 260 mg/l cytidine and 500 mg/l uridine, uninucleate microspores developed into mature pollen at a 12% germination rate. Our report is the first demonstration of maturation of S. nigra microspores in vitro.     

响应面法优化麻疯树花粉离体萌发培养基的研究

麻疯树因其种子含油率较高,种子油提炼的生物柴油可部分替代汽油,而成为一种极具潜能的能源作物,但由于产量低,麻疯树在热带、亚热带的发展受到极大限制。杂交育种是提高产量的重要手段,杂交亲本花粉生活力的高低直接影响到育种的成效。因此,寻求麻疯树离体花粉萌发的最适培养基配方,探明花粉萌发培养基中各主要培养基成分间的交互作用对生产上麻疯树杂交结实率和种子产量的提高具有重要意义。该研究以麻疯树开花初期雄花上花药刚散粉时的成熟花粉粒为材料,采用Box-Behnken设计(Box-Behnken design,BBD)的响应面法,对麻疯树花粉离体萌发培养基中各主要培养基成分的浓度配比及各主要培养基成分的交互作用进行了研究。以花粉萌发率为响应指标,建立了4种营养成分(蔗糖、硼酸、硝酸钙、硝酸钾)与花粉萌发率的响应面模型,并对各主要培养基成分的浓度配比进行了优化。通过R软件进行响应面分析的结果表明:4因素对花粉萌发率的影响顺序为蔗糖硼酸硝酸钙硝酸钾;蔗糖与硼酸、蔗糖与硝酸钙、蔗糖与硝酸钾之间的交互作用显著。响应面建模优化后的最佳培养基为13.77%蔗糖+32.14 mg·L~(-1)硼酸+22.21 mg·L~(-1)硝酸钙+19.95 mg·L~(-1)硝酸钾+200 mg·L~(-1)硫酸镁,在此条件下的理论萌发率为99.73%。采用此培养基成分配比得到麻疯树花粉离体试验萌发率为98.97%,与理论响应值相吻合,同时也表明利用BBD设计的响应面模型进行麻疯树花粉离体萌发培养条件优化方法的有效性。     

The vitality of the Serbian spruce (Picea omorika) pollen during the long-term cryopreservation

Abstract

This article studies differences in the vitality (germination rate and pollen tube length) of fresh pollen and pollen stored at ?20 °C for 14 years of Serbian spruce (Picea omorika [Pan?i?] Purkyne). Fresh pollen was sampled from 24 trees from a Serbian spruce seed stand area on the site of Bela zemlja (43° 48′ 32″ N, 19° 44′ 25″ E) in the area of Mount Zlatibor (Serbia) in two successive years. Six nutrition media with different sucrose concentrations (0%, 5%, 10%, 15%, 20% and 25% aqueous solution) were used to assess the vitality of the pollen. Variance analysis shows significant differences among years of pollen sampling, years of cryopreservation, trees, and sucrose treatments for pollen germination for both germination rate and pollen tube length. The results indicate that climate conditions preceding the pollen maturation period had an impact on the pollen quality as well as on the pollen vitality during the years of cryopreservation. Inter-individual variability (differences between trees genetic variability) was very pronounced and enabled selection of pollinator trees that retained their initial vitality during the period of cryopreservation. This periodicity in Serbian spruce full flowering might have significant practical application in the breeding of this important species.     

The division of the generative nucleus and the formation of callose plugs in pollen tubes of Aechmea fasciata (Bromeliaceae) cultured in vitro

The effect of different external factors on pollen germination and pollen tube growth is well documented for several species. On the other hand the consequences of these factors on the division of the generative nucleus and the formation of callose plugs are less known. In this study we report the effect of medium pH, 2-[N-morpholino]ethanesulfonic acid (MES) buffer, sucrose concentration, partial substitution of sucrose by polyethyleneglycol (PEG) 6000, arginine (Arg), and pollen density on the following parameters: pollen germination, pollen tube length, division of the generative nucleus, and the formation of callose plugs. We also studied the different developmental processes in relation to time. The optimal pH for all parameters tested was 6.7. In particular, the division of the generative nucleus and callose plug deposition were inhibited at lower pH values. MES buffer had a toxic effect; both pollen germination and pollen tube length were lowered. MES buffer also influenced migration of the male germ unit (MGU), the second mitotic division, and the formation of callose plugs. A sucrose concentration of 10% was optimal for pollen germination, pollen tube growth rate and final pollen tube length, as well as for division of the generative nucleus and the production of callose plugs. Partial substitution of sucrose by PEG 6000 had no influence on pollen germination and pollen tube length. However, in these pollen tubes the MGU often did not migrate and no callose plugs were observed. Pollen tube growth was independent of the migration of the MGU and the deposition of callose plugs. In previous experiments Arg proved to be positive for the division of the generative nucleus in pollen tubes cultured in vitro. Here, we found that more pollen tubes had callose plugs and more callose plugs per pollen tube were produced on medium with Arg. After the MGU migrated into the pollen tube (1 h after cultivation), callose plugs were deposited (3 h). After 8 h the first sperm cells were produced. The MGU moved away from the active pollen tube tip until the second pollen mitosis occurred, thereafter the distance from the MGU to the pollen tube tip diminished. Callose plug deposition never started prior to MGU migration into the pollen tube. Pollen tubes without a MGU also lack callose plugs (±30% of the total number of pollen tubes). Furthermore, we found a correlation between the occurrence of sperm cells in pollen tubes and the synthesis of callose plugs.     

Effect of enhanced UV-B radiation on pollen quantity,quality, and seed yield in Brassica rapa (Brassicaceae)

We conducted three experiments to examine the influence of ultraviolet-B radiation (UV-B; 280–320 nm) exposure on reproduction in Brassica rapa (Brassicaceae). Plants were grown in a greenhouse under three biologically effective UV-B levels that simulated either an ambient stratospheric ozone level (control), 16% (“low enhanced”), or 32% (“high enhanced”) ozone depletion levels at Morgantown, WV, USA in mid-March. In the first experiment, we examined whether UV-B level during plant growth influenced in vivo pollen production and viability, and flower production. Pollen production and viability per flower were reduced by ≈50% under both enhanced UV-B levels relative to ambient controls. While plants under high-enhanced UV-B produced over 40% more flowers than plants under the two lower UV-B treatments, whole-plant production of viable pollen was reduced under high-enhanced UV-B to 17% of that of ambient controls. Whole-plant production of viable pollen was reduced under low-enhanced UV-B to 34% of ambient controls. In the second experiment, we collected pollen from plants under the three UV-B levels and examined whether source-plant UV-B exposure influenced in vitro pollen germination and viability. Pollen from plants under both enhanced-UV-B treatments had initially lower germination and viability than pollen from the ambient level. After in vitro exposure to the high-enhanced UV-B levels for 6 h, viability of the pollen from plants grown under ambient UV-B was reduced from 65 to 18%. In contrast, viability of the pollen from plants grown under both enhanced UV-B treatments was reduced to a much lesser extent: only from ≈43 to 22%. Thus, ambient source-plant pollen was more sensitive to enhanced UV-B exposure. In the third experiment, we used pollen collected from source plants under the three UV-B levels to fertilize plants growing under ambient-UV-B levels, and assessed subsequent seed production and germination. Seed abortion rates were higher in plants pollinated with pollen from the enhanced UV-B treatments, than from ambient UV-B. Despite this, seed yield (number and mass) per plant was similar, regardless of the UV-B exposure of their pollen source. Our findings demonstrate that enhanced UV-B levels associated with springtime ozone depletion events have the capacity to substantially reduce viable pollen production, and could ultimately reduce reproductive success of B. rapa.     

梅花花粉离体萌发和花粉管生长研究

(南京农业大学园艺学院,南京210095)摘要:研究培养基成分、pH值和培养方式对梅花花粉离体萌发和花粉管生长的影响。结果表明:不同品种梅花花粉离体萌发的最适培养基为ME3+200g.L-1PEG4000(pH5.0),品种‘淡丰后’、‘久观绿萼’、‘喧妍宫粉’和‘月光玉蝶’最高萌发率可分别达到58.6%、60.6%、85.6%和50.7%。PEG4000能显著促进梅花花粉萌发,在培养基各成分中作用最大,不可替代。低浓度(50g.L-1)蔗糖对梅花品种花粉萌发作用不显著,而高浓度(≥100g.L-1)蔗糖明显抑制花粉萌发和花粉管生长。固体和液体培养对梅花花粉离体萌发的影响差异不显著。     

Germination of trinucleated pollen: formulation of a new medium for Capsella bursa-pastoris

Summary In Brewbaker and Kwack's medium (BK) only 16% of the pollen grains germinated, and these produced pollen tubes having a maximum length of 25 m. With a solution based on Monnier's medium 47% germination and 160-mlong pollen tubes were observed. Calcium was shown to be essential for germination; the optimal concentration was 880 mg/l calcium chloride. The optimal concentrations of magnesium sulphate and boric acid were 360 and 50 mg/l, respectively. Germination at pH 4.0 but also pH 8.0 and the presence of vitamins B1 and B6 (1 mg/l each) were stimulatory. Polyethylene glycol (PEG) was superior to sucrose as an osmoticum and germination and tube length were significantly improved using PEG 4000 at a concentration of 120 g/l (0.03 M). Equimolar concentrations of PEG 400 and PEG 600 gave inferior results. Combining PEG with sucrose in the medium did not improve germination or increase tube length.     

桃花粉离体萌发和花粉管生长特性研究

采用花粉离体萌发法研究不同培养基组分和培养条件对桃花粉萌发和花粉管生长的影响,同时对不同贮藏温度下的桃花粉寿命进行研究.结果表明:固体培养基与液体培养基对桃的花粉萌发率和花粉管长度影响差异不显著;10%蔗糖是大多数桃品种花粉的最适萌发条件;硼能提高桃花粉的萌发率,但对花粉管的生长没有促进作用;桃花粉在20℃～25℃的培养温度下萌发率最高,花粉管最长;桃花粉萌发率和花粉管长度在培养前3 h内上升最快,3～5 h上升趋势减弱,5 h后基本停止;随着贮藏温度的升高和贮藏时间的延长,花粉生活力呈降低的趋势.     

Pollen germination and pollen tube growth in Fraxinus pennsylvanica

With regard to adaptation of green ash (Fraxinus pennsylvanica Marshall) to ecological conditions in Croatia, pollen germination and pollen tube length after 2, 4 and 6 hours were examined in vitro at 10, 15, 20 and 25°C during two years 2001 and 2002. Narrow leaved ash (F. angustifolia Vahl) pollen served as a control in 2002. The year, time and temperature, and the interaction between time and temperature were significant for both germination percentage and pollen tube length. Interactions year × temperature and year × time were significant for pollen tube length only. The highest germination percentage (17.86% in 2001 and 19.40% in 2002) of green ash pollen was at 15°C after 6 hours. The pollen tube length was greatest at 20°C (393.46 μm) in 2001 and 25°C (899.50 μm) in 2002 after 6 hours. Narrow leaved ash pollen had the highest germination percentage (19.22%) at 20°C after 6 hours and was significantly reduced at 25°C. The pollen tube length was greatest at 25°C (518.90 μm) after 6 hours. It can be concluded that green ash pollen has satisfactory germination in ecological conditions in Croatia and that the optimum temperature for pollen germination is higher than 20°C.     

Observation of Flowering Process of Grape (<i>Vitis vinifera</i> L.)—Insight into the Starting of Pollination of Flower Hiding in Calyptras (Cap)

It is generally agreed that many Vitis vinifera L. cultivars are self-fertile, where self-pollination often occurs before capfall in a process called cleistogamy. Therefore, it is difficult to identify the right time to remove stamens before self-pollination during the cross-breeding of grape. For this paper, we observed the process of grape flowering and measured the pollen viability and stigma receptivity of grape flowers of ‘Shine Muscat’ in order to identify the starting time of self-pollination before capfall and to provide useful information for improving the efficiency of cross-breeding. The results demonstrate that the anther is not cracked during the visible clusters and separated clusters stages. Meanwhile, in the separated floral buds, flowering begins, and full bloom stages, the pollen viability is 60.7%, 73.2% and 80.3%, respectively; however, at the berry set stage, pollen viability drops to zero. The top of the mature stigma is composed of a layer of nearly cylindrical papillary cells, and the stigma receptivity for pollen changes with the development of flowers: in particular, no reaction was observed in the visible clusters stage; weak positive reaction at the separated clusters stage; strong positive reaction at the separated floral buds, flowering begins, and full bloom stages; and no reaction at the berry set stage. In the separated floral buds stage, pollen tubes were seen germinating in the style. In the flowering begins stage, more pollen tubes were observed at the entry of the ovary. During the full bloom stage, most pollen tubes elongated into the ovary base and some entered the pearl hole. At the berry set stage, newborn endosperm nucleus could be seen in the ovule. From the above, we can conclude that the initiation time of closed fertilization for ‘Shine Muscat’ grape can be judged as the separated floral buds stage, and it is best to discard the stamen before the separated floral buds stage when conducting cross-breeding.     

Division of the generative nucleus in cultured pollen tubes of the Bromeliaceae

Pollen germination, division of the generative nucleus and position of the generative nucleus in the pollen tube during in vitro germination were examined for six bromeliad cultivars. The influence of mixed amino acids (casein hydrolysate) and individual amino acids (Arg, Asn, Asp, Glu, Gly, Met, Phe, Orn, Tyr) were tested. Aechmea fasciata and A. chantinii pollen tubes showed more generative nuclear division in cultured pollen tubes than the other four cultivars tested. Casein hydrolysate did not stimulate generative nuclear division. In general arginine (1 mM) improved division of the Aechmea generative nucleus and to a lesser extent this of Vriesea `Christiane', Guzmania lingulata and Tillandsia cyanea. A concentration of 2 mM arginine reduced pollen tube growth of Aechmea. The vegetative nucleus was ahead of the generative nucleus in approximately 50% of the pollen tubes of all cultivars studied. In about 25% of the pollen tubes, the generative nucleus was ahead and in ±25% pollen tubes the vegetative and generative nuclei were joined together. The distance between the two generative nuclei and the distance from the generative nuclei to the pollen tube tip differed significantly for Aechmea fasciata and A. chantinii. The influence of different amino acids for Aechmea fasciata and A. chantinii varied with respect to pollen germination and generative nuclear division. Arg and Met improved nuclear division of both Aechmea cultivars. Pollen germination and sperm cell production were not linked. This information is important to ameliorate in vitro pollination methods used to overcome fertilization barriers in Bromeliaceae and other higher plants.     

Isolation of Allium pollen protoplasts

Studies on protoplasts isolation were carried out with mature pollen grains of 29 samples of species of Allium aflatunense, A. cepa, A. fistulosum, A. karataviense, A. longicuspis, A. nutans, A. odorum, A. sativum and A. schoenoprasum. Surface sterilized pollen grains drifted from crushed anthers were incubated in an enzyme solution containing 1% (w/v) cellulase Onozuka R-10, 1% (w/v) Macerozyme R-10, 0,5 mol l^-1 sucrose and the basal salts of Nitsch medium. Protoplasts were released within 3 to 120 min, either from the pollen grain, through a slightly disturbed germination pore (narrow aperture), or through a wider aperture, when the exine surrounding the germination pore was disturbed. For the first time, protoplasts were obtained from 13 genotypes of 6 Allium species, at a rate of 1 to 30% of the digested intact pollen grains, depending on the genotype.     

Forage quality and quantity affect red mason bees and honeybees differently in flowers of strawberry varieties

Nectar is a vital source of energy for bees and other pollinators and pollen represents the only source of protein in the diet of bees. Nectar and pollen quality and quantity can therefore affect foraging choices. Strawberry, Fragaria × ananassa (Rosaceae), is a flowering crop that requires insect pollination for the berries to develop optimally. The solitary red mason bee, Osmia bicornis L. (Hymenoptera: Megachilidae), occurs naturally but like the eusocial western honeybee, Apis mellifera mellifera L. (Hymenoptera: Apidae), it is also a commercially reared pollinator used in strawberry production. We hypothesized that strawberry nectar and pollen quality would affect the foraging choice of these two types of bees. In this study nectar and pollen quality is represented by various levels of sugar and protein content, respectively, as well as the number of open strawberry flowers in the experimental field, would affect the foraging choice of these two types of bees. Consistent with previous studies, we found significant and major differences between strawberry varieties in proportions of sucrose in the nectar sugar and in pollen viability – a proxy for pollen protein content. All measured parameters had a significant effect on red mason bee visitation frequency. Contrary to expectations, honeybee foraging behavior was only affected by the number of open flowers and not by any of the quality parameters measured. Our findings indicate that red mason bees were capable of assessing nectar and pollen quality and prioritize accordingly. The pattern observed indicates that individual red mason bees changed foraging focus between strawberry varieties depending on whether nectar or pollen was collected. Our results suggest that targeted breeding of varieties toward high levels of nectar sugar and sucrose concentrations and high pollen protein content may increase pollination success from red mason bees and possibly other solitary bees.     

Influence of arginine,ornithine, DFMO and polyamines on division of the generative nucleus in cultured pollen tubes of <Emphasis Type="Italic">Aechmea fasciata</Emphasis> (Bromeliaceae)

During in vitro pollen tube growth of Aechmea fasciata the second pollen mitosis (PM II) that produces two sperm cells was influenced by exogenous amino acids. Arginine (Arg) as single amino acid was the limiting factor for the second mitosis of the generative nucleus and thus the formation of sperm cells in cultured pollen tubes of A. fasciata. The involvement of Arg was probably related to protein synthesis. The need for Arg was not related to polyamine (PA) biosynthesis, since PA added to the germination medium were unfavourable for sperm cell production. Both ornithine (Orn) and difluoromethylornithine (DFMO) inhibited the second mitosis in cultured pollen tubes of A. fasciata. The addition of Arg during the first 2 h of pollen germination was necessary to establish the division of the generative nucleus 6 h later.     

The Germination of Vinca rosea L. Pollen Grains and the Growth of the Pollen Tubes in vitro

The effect of medium concentration, pollen grain concentration, pH of the media, light and temperature on the germination of Vin ca rosea pollen grains, and the growth of their pollen tubes in vitro have been studied. The pollen grains germinate best at a sucrose concentration between 14.2% and 30%; when the pollen grain concentration exceeds 800 per 0.0234 ml; at near neutral pH (6.5); in darkness and at a temperature close to 30°. Moreover buffering ions affect the growth of the pollen tubes. Pollen grains remain viable in a wide range of temperatures, and the wall of the pollen grain is capable of withstanding severe osmotic imbalance. Low temperature induces spherical swellings at the tips of the pollen tubes, followed by accumulation of a hyaline plug.     

七叶树花粉活力和柱头可授性变化的研究

采用花粉离体萌发法研究不同蔗糖浓度、硼酸浓度和不同温度对七叶树花粉萌发的影响及花粉活力测定,用联苯胺-过氧化氢法测定柱头可授性.结果表明:七叶树花粉萌发的最佳培养基是12％蔗糖+30 mg/L硼酸,花粉萌发的最适温度为25℃.雄花在开花当天花粉活力最高达75.69％,并在开花当天的上午10:00时,花粉活力最强,10:00～16:00花粉活力保持较高活力,是最佳授粉时段.两性花的柱头可授期持续时间较长,为8～9 d,开花3d达到最强,开花1～4 d柱头可授性保持较高水平,为授粉的最佳时间段.因此,从七叶树的花部特征、花粉活力与柱头可授性及花粉萌发的条件看,在长期的自然选择下七叶树在花部结构和开花生理上都是相配合的,以保障生殖成功.     

山玉兰花粉离体萌发和花粉管生长特性的研究

山玉兰(Magnolia delavayi)是木兰科木兰属的常绿乔木或大型灌木,是重要的园林造景、庭院绿化素材,也是重要的育种资源。山玉兰花粉的研究对其杂交育种的成败具有重要影响,但目前尚未见其花粉活力的相关报道。该研究以新鲜的山玉兰花粉为对象,采用悬滴培养法分析了温度、光照以及培养液的蔗糖和硼酸浓度对山玉兰花粉萌发的影响。结果表明:(1)山玉兰花粉萌发时,最适宜的温度为27℃。(2)光暗条件下,山玉兰花粉以浓度为5%的蔗糖培养效果最佳,其萌发率在16%以上;而硼酸浓度则以0.001%的培养效果最佳。(3)蔗糖与硼酸共同作用可有效促进花粉萌发和花粉管生长。其中,在光照条件下,以5.0%蔗糖+0.001%硼酸为最适宜的培养液,花粉萌发率达41.27%,花粉管长达281.49μm;而在黑暗条件下,则以5.0%蔗糖+0.01%硼酸为最适宜的培养液,花粉萌发率达45.71%,花粉管长达254.00μm。该研究结果为进一步开展人工辅助授粉、发掘山玉兰的种质资源工作奠定了基础。     

The reversible inhibition of pollen germination of Nicotiana tabacum L.: an entry into a transformation system

Summary The hydrodynamics of mature pollen rehydration in Nicotiana tabacum was used to study reversible inhibition of pollen germination in vitro. Tobacco pollen was incubated for various times in media containing calcium, potassium and magnesium salts, boric acid, and exhibiting different osmotic pressures as a function of sucrose concentration. Total inhibition of germination with complete viability preservation was achieved for 56 h by keeping the grains in medium with 80% sucrose, since typical percentages of germination and pollen tube lengths were recovered after this treatment. These effects were considered as consequences of natural osmoregulation of rehydration/germination in mature pollen. The possibility of applying these findings to the incubation of pollen with Agrobacterium tumefaciens to develop a pollen transformation method is discussed.     

Effect of smoke derivatives on in vitro pollen germination and pollen tube elongation of species from different plant families

Plant‐derived smoke stimulates seed germination in numerous plant species. Smoke also has a positive stimulatory effect on pollen germination and pollen tube growth. The range of plant families affected my smoke still needs to be established since the initial study was restricted to only three species from the Amaryllidaceae. The effects of smoke‐water (SW) and the smoke‐derived compounds, karrikinolide (KAR₁) and trimethylbutenolide (TMB) on pollen growth characteristics were evaluated in seven different plant families. Smoke‐water (1:1000 and 1:2000 v:v) combined with either Brewbaker and Kwack's (BWK) medium or sucrose and boric acid (SB) medium significantly improved pollen germination and pollen tube growth in Aloe maculata All., Kniphofia uvaria Oken, Lachenalia aloides (L.f.) Engl. var. aloides and Tulbaghia simmleri P. Beauv. Karrikinolide (10^?6 and 10^?7 m ) treatment significantly improved pollen tube growth in A. maculata, K. uvaria, L. aloides and Nematanthus crassifolius (Schott) Wiehle compared to the controls. BWK or SB medium containing TMB (10^?3 m ) produced significantly longer pollen tubes in A. maculata, K. uvaria and N. crassifolius. These results indicate that plant‐derived smoke and the smoke‐isolated compounds may stimulate pollen growth in a wide range of plant species.     

Disentangling the components of pollen limitation in a widespread herb with gametophytic self-incompatibility

Premise

Seed production is frequently limited by the receipt of insufficient or low-quality pollen, collectively termed “pollen limitation” (PL). In taxa with gametophytic self-incompatibility (GSI), incompatible pollen can germinate on stigmas but pollen tubes are arrested in styles. This allows for estimates of pollen performance before, during, and after self-recognition, as well as insight into the factors underlying pollen quality limitation in GSI taxa.

Methods

We scored pollen performance following self and outcross pollinations in Argentina anserina to identify the location of self-recognition and establish the relationship between pollen tubes and seed production. We then estimated quantity and quality components of PL from >3300 field-collected styles. We combined our results with other studies to test the prediction that low pollen quality, but not quantity, drives higher PL in self-incompatible (SI) taxa than in self-compatible taxa (SC).

Results

Self and outcross pollen germinated readily on stigmas, but 96% of germinated self-pollen was arrested during early tube elongation. Reproduction in the field was more limited by pollen quality than by quantity, and pollen failure near the location of self-recognition was a stronger barrier to fertilization than pollen germination. Across 26 taxa, SI species experienced stronger pollen quality, but not quantity, limitation than SC species.

Conclusions

Evaluating pollen performance at multiple points within pistils can elucidate potential causes of pollen quality limitation. The receipt of incompatible pollen inhibits fertilization success more than insufficient pollen receipt or poor pollen germination in A. anserina. Likewise, pollen quality limitation drives high overall PL in other SI taxa.